Mercury and most of its derivatives are extremely toxic substances which are ubiquitous in the biosphere. The major source of mercury contamination is the natural degassing of the earth's crust, although major contributions also arise from anthropogenic sources. The amount released from both sources do not cause problems on a global scale, but increases on a local level can lead to serious health problems with long-term consequences for the affected population. Thus, simple, sensitive and reliable procedures for the detection of mercury in the environment are needed.
Numerous approaches for detecting transition/heavy metals with sensitivity in the parts per million (ppm)/high parts per billion (ppb) range can be found in the literature (see, e.g., Blanco, M., et al., Mikrochira. Acta, 108: 53-59 (1992)). These approaches are primarily based on the detection of suitable metal derivatives by molecular absorption spectrophotometry (e.g., UV-VIS, IR) or by electroanalytical techniques. Many of these techniques employ the use of chelates derived from the metal ions present in the sample being analyzed. Metal ions in complex matrices have also been measured in the form of chelates by both gas chromatography (GC) and liquid chromatography (LC).
Simple mercury analyses based on the detection of chelates using optical techniques (such as, for example, UV-VIS photometry) often suffer from interferences by foreign ions (see, Sharma, R., et at., Talanta, 36: 457-461 (1989)), and they usually lack the necessary high sensitivity (see, Madseal, M., J. Anal. Chem., 342: 157-162 (1992); and Kamburova, M., Talanta 40(5): 719-723 (1993). This is true despite the fact that these methods are frequently combined with chelate extraction techniques (e.g., preconcentration techniques). Only occasionally have detection limits below 0.1 ppm been reported in the literature (see, Mariscal, M., et al., ibid.).
As such, relatively few simple, reliable, and selective techniques are available for the detection of transition/heavy metals at parts per billion (ppb)/parts per trillion (ppt) levels. The most commonly used methods involve instrumental analyses, such as, for example, flameless atom absorption spectroscopy, inductively coupled plasma mass spectrophotometry, special electroanalytical methods, etc. (See, Daniels, R., et al., Sci. Total Environ., 89: 319-339 (1989).) These methods, however, possess many of the limitations that are typically encountered when using instrumental analyses: the sample throughput is limited; they cannot process multiple samples in parallel; and they are not suitable or amendable to on-site analysis in the field (see, Wylie, D., et al., Anal. Biochem., 194: 381-387 (1991)).
Interesting alternative methods involve immunoassay procedures for the detection of certain lanthanida elements and heavy metals (see, Reardan, et al., Nature, 316: 265-268 (1985); Wylie, D., et al., ibid., (1991); Wylie, D., et al., Proc. Natl. Acad. Sci. USA, 89: 4104-4108 (1992)). In these methods, antibodies are raised against metal chelates. Reardan, et al. succeeded in the recognition of an EDTA-type of chelate of indium by monoclonal antibodies generated by the same chelate attached to a carrier protein. The antibodies displayed the highest affinity to the indium-chelate; however, chelates of various heavy metal ions had affinities of only 10-1000 times less than the indium-chelate. This lack of selectivity can create significant difficulties if real samples containing other metal ions in moderate to high concentrations were to be analyzed using this method. Moreover, Wylie, et al., ibid., immunized mice with HgCl.sub.2 -glutathione-KLH. Some of the monoclonal antibodies that were obtained selectively recognized the mercuric ions either chelated to glutathione-BSA or alone. One of the resulting ELISAs, based on the use of the most effective antibodies, showed a linear relationship between the optical density and log[Hg.sup.2+ ] in the range of 0.5-10 ppb. Although these method exhibit some degree of specificity, they both have significant limitations in that they require the use of highly specific monoclonal antibodies that are difficult and expensive to prepare.
As such, in environmental and clinical laboratories, the most routinely used method for the determination of mercury is cold-vapor atomic absorption spectrometry (i.e., CVAAS). (See, Magos, L., et at., J. Assoc. Of Anal. Chem., 55: 966-971 (1972); Lind, et al., Fresenius J. Anal. Chem. 345: 314-317 (1993); Guo, T., et al., J. Anal. Chim. Acta., 278: 189-196 (1993)). However, this technique also has a number of limitations. As with the other methods involving instrumental analyses, the number of samples that can be analyzed is limited by the fact that only one sample can be analyzed at a time. Additionally, a large sample volume (i.e., up to 5 mL) is required to ensure maximum sensitivity, although assays performed at maximum sensitivity often lack high precision. Moreover, CVAAS requires the use of expensive equipment and highly skilled personnel, and it is not suitable or amendable to on-site analysis in the field (see, Wylie, et al., ibid. (1991)).
In the case of mercury, chemical speciation (i.e., inorganic versus organic mercury) is of utmost toxicological importance. Of particular interest is methylmercury. Methylmercury, produced by bacterial methylation of Hg(II) in aquatic sediments, is by far the most toxic and most commonly occurring organic mercury species (see, Palmisano, F., et al., J. Anal. Chem., 346: 648-652 (1993)). It is significantly more toxic than Hg(II), i.e., inorganic mercury. Methylmercury accumulates in fish and is amplified through the food chain; thus, its concentration in natural waters is one of the most important water quality parameters (see, Bloom, N., et al., Water Air Soil Pollut., 53: 251-265 (1990)). The abundance and the toxicological significance of Hg(0) and Hg(I) in aquatic environments is only marginal. The CVAAS technique detects the amount of Hg(II), i.e., inorganic mercury. However, the quantification of organic mercury (i.e., total mercury-inorganic mercury) is possible indirectly by decomposition of the organic species into Hg(II) and then by the determination of the total mercury content (see, Magos, L., ibid.; Lind, et al., ibid.; Guo, T., et al., ibid.). Other common methods for determining mercury speciation combine chromatographical separation of the mercury derivatives using various detection systems (see, Bulska, E., et al., Analyst, 117: 657-663 (1992); Palmisano, F., et al., ibid.; Lind, et al., ibid.). However, all of these methods require the use of costly apparatuses and highly qualified analysts. Moreover, the analysis rate is limited because only one sample can be measured at a time.
As such, there still exists a need for a simple, sensitive and reliable methods for the detection of mercury and other metal ions in environmental and biological samples. The present invention remedies this need by providing such methods.